Asthma is a complex, chronic disorder, with a genetic and an environmental component (1). It is characterized by reversible airway obstruction, airway hyperresponsiveness, airway inflammation and remodeling (2). Asthma affects an estimated 15 million Americans and the morbidity and mortality associated with it is on the rise in industrialized countries (3,4). Inflammation in the airway of an allergic asthmatic is associated with the mucosal infiltration of T helper (Th)2 subset of CD4+ T cells and eosinophils (5,6). The interaction between these cells leads to the production of various pro-inflammatory mediators involved in the pathogenesis of asthma (7,8). Other forms of asthma are those that are induced by exercise, viruses, aspirin and occupation. Although the mechanism responsible for these forms of asthma might involve Th2 lymphocytes and cytokines it might be triggered differently (9-12). Many cytokines and chemokines are involved in the pathogenesis of asthma (13,14). Specifically, the Th2 derived cytokines (interleukin 4, 5, 9 and 13) play an important role in allergic diseases including asthma.
Chronic obstructive pulmonary disease (COPD) is a chronic pulmonary inflammation characterized by the infiltration of neutrophils, macrophages, B and T cells. These immunocompetent cells are activated by a variety of cytokines and chemokines that are released in the lung in response to a prolonged exposure to toxic gases and particles (15). Bronchitis and emphysema, together with irreversible airflow obstruction, are the clinical manifestations of the disease. No known agents delay the accelerated decline in pulmonary function that characterizes COPD.
Recently, it was observed that the progression of COPD was strongly associated with the parenchymal infiltration by innate and adaptive inflammatory immune cells forming an ectopic lymphoid follicle containing a germinal center. The presence of the lymphoid follicle was coupled to a remodeling process that thickened the distal small airway walls (16). This result strongly suggests the potential pathological role of the ectopic lymphoid follicles in COPD.
In an effort to identify novel genes involved in the pathogenesis of asthma, researchers have used DNA microarray technology to profile genes that are differentially expressed in animal models of asthma (17,18). Microarray technology is a powerful tool since it enables analysis of the expression of thousands of genes simultaneously and can also be automated allowing for a high-throughput format. In multifactorial diseases, such as asthma, microarray results can provide a gene expression profile which can prove very useful in designing new therapeutics. Also, it can prove very powerful in identifying novel genes and annotating genes of unknown function (19).
CXCL13 (a.k.a BLC (B cell homing chemokine) or BCA-1 (B cell attracting chemokine 1) or Angie 2)) is a chemotactic factor that most strongly and selectively attracts B cells. It also promotes migration of certain T cells and macrophages through the receptor CXCR5 (20). CXCL13 is expressed in the follicles of Peyer's patches, spleen and lymph nodes, and is believed to be important in follicle development and homeostasis (21).
It has been observed for years that in the sites of chronic inflammation, the arrangement of the inflammatory infiltrate (T, B and stromal cells) shares many architectural features with lymphoid tissue, which forms the so called ectopic lymphoid follicles (21). In addition, extopic high production of CXCL13 is associated with lymphocyte accumulation and ectopic lymphoid follicle formation in chronic inflammatory diseases, such as rheumatoid arthritis (21), Sjogren's syndrome (22), various forms of lupus such as systemic lupus erythematosus (23, 24), ulcerative colitis (25, 26), multiple sclerosis (27-29), type I diabetes (30-32) and autoimmune thyroid diseases (33, 34). Although the exact pathogenic role of the ectopic lymphoid follicles is not clear, evidence suggests its importance in the switch from acute, resolving to chronic, persistent inflammation by allowing lymphocytes to accumulate in the local inflamed tissue (35). Therefore, disrupting or eliminating the ectopic lymphoid follicles would provide a novel therapeutic approach to control chronic inflammatory diseases. CXCL13 is an ideal therapeutic target due to its high expression levels in the ectopic lymphoid follicles and its role in maintaining their microstructure and attracting B cells.
Systemic lupus erythematosus (SLE or lupus) is a chronic autoimmune disease that is potentially debilitating and sometimes fatal as the immune system attacks the body's cells and tissue, resulting in inflammation and tissue damage. SLE can affect any part of the body, but most often harms the heart, joints, skin, lungs, blood vessels, liver, kidneys and nervous system.
The CXCL13 gene (GenBank Accession No. NM—006419, SEQ ID NO:1) resides on human chromosome 4q21. CXCL13 belongs to the CXC chemokine family. CXCL13 is critical for lyphoid organ formation/development, B cell follicle formation, and B cell recruitment. It is highly produced ectopically in the inflamed tissues of multiple chronic inflammatory diseases, and is believed to play an important role in maintaining local B and T cell activation and inflammation.
Gene expression can be modulated in several different ways, including by the use of siRNAs, shRNAs, antisense molecules and DNAzymes. SiRNAs and shRNAs both work via the RNAi pathway and have been successfully used to suppress the expression of genes. RNAi was first discovered in worms and the phenomenon of gene silencing related to dsRNA was first reported in plants by Fire and Mello and is thought to be a way for plant cells to combat infection with RNA viruses. In this pathway, the long dsRNA viral product is processed into smaller fragments of 21-25 bp in length by a DICER-like enzyme and then the double-stranded molecule is unwound and loaded into the RNA induced silencing complex (RISC). A similar pathway has been identified in mammalian cells with the notable difference that the dsRNA molecules must be smaller than 30 bp in length in order to avoid the induction of the so-called interferon response, which is not gene specific and leads to the global shut down of protein synthesis in the cell.
Synthetic siRNAs can be designed to specifically target one gene and they can easily be delivered to cells in vitro or in vivo. ShRNAs are the DNA equivalents of siRNA molecules and have the advantage of being incorporated into the cells' genome and then being replicated during every mitotic cycle.
DNAzymes have also been used to modulate gene expression. DNAzymes are catalytic DNA molecules that cleave single-stranded RNA. They are highly selective for the target RNA sequence and as such can be used to down-regulate specific genes through targeting of the messenger RNA.
Accordingly, there is a need to identify and characterize new methods for diagnosing and treatment related to CXCL13 for pulmonary disorders, such as asthma, and related diseases and conditions. Additionally, there is a need to identify and characterize new methods for treating disorders such as systemic lupus erythematosus.